This invention is directed to sequencing of polypeptides and proteins.
Biotechnologists have identified proteins in animal, plant and microorganism cells that combat or prevent disease, confer resistance to plants for insects, improve the taste, appearance and shelf life of agricultural products, prevent fouling of hulls of boats, and can be used to clean soiled clothing. These proteins are made of amino acids linked together. There are 20 commonly occurring amino acids. The order in which the amino acids are linked together is known as the primary structure of the protein. A short chain of amino acids is called a polypeptide.
Once a protein is identified, it can be mass produced and put to constructive use. To do this, the protein needs to be sequenced, i.e., determine which amino acids constitute the protein and in what order they are linked together. The sequence data is then used in conjunction with the genetic code to prepare probes to find the gene within the cell which codes for that protein. Once the gene is identified, it can be cloned into a suitable host cell for expressing the gene and producing the protein.
Sequencing of proteins can also be useful for gene therapy or alteration. In gene therapy or alteration, a protein of interest is identified within a cell. The protein is then sequenced, and the sequence information is used in conjunction with the genetic code to make probes to identify the gene which codes for the protein. Once the gene is identified, it may be possible to manipulate that gene in an animal or plant to turn it "on" or "off", replace it with another gene, or transfer it to another animal or plant.
The process of sequencing a macromolecule such as a protein or polypeptide has been automated. There are two basic approaches for manipulating a macromolecule in the automated sequencing operation. One approach is to bond the macromolecule to a substrate such as support beads and pack the support beads into a column. This approach suffers from the disadvantage that the support beads can swell or become fouled, resulting in the column becoming plugged. Also, the beads can shrink resulting in undesirable channeling.
The second sequencing process is to bond the macromolecule to a substrate such as filter paper or polymeric membranes. This approach suffers from the disadvantage of the lack of any built-in system for determining the accuracy of the sequencing technique. There is no mechanism available to determine if the sequencing process is performing properly while actually doing the sequencing.
Accordingly, there is a need for a sequencing process which overcomes problems related to bead shrinkage, bead swelling, and column plugging; and includes a built-in system for determining the accuracy of the sequencing results.